与人体肥胖相关的神经肽Y受体Y2基因的克隆及序列分析

Cloning and sequence analysis of the human obesity- related neuropeptide Y（NPY） Y2 receptor gene

目的克隆与肥胖相关的人神经肽Y受体Y2（NPY2R）基因，并鉴定该克隆基因序列的正确性。方法从人的脂肪组织中提取总RNA并进行反转录，以此为模板用PCR扩增NPY2R基因的eDNA，然后与pET28a＋载体重组，筛选阳性克隆和DNA序列分析鉴定，测序后与GeneBank中NPY2R基因进行同源性比较和序列分析。结果PCR扩增出一个1100bp左右的DNA片段，与载体重组和DNA序列分析鉴定，DNA序列分析显示克隆的DNA片段是人NPY2R基因，且所克隆的基因共编码381个氨基酸，分子量为4．2kD，与GeneBank中NPY2R基因序列同源性达100％。结论克隆的人NPY2R基因与GeneBank中NPY2R序列完全一致，为进一步应用分子生物学技术深入研究NPY2R与人体肥胖发生、发展及转化等相关作用机制及应用该基因进行其基因蛋白表达奠定了基础。

Objective The present paper aims at cloning and identifying human neuropeptide Y receptor Y2 （ NPY2R）, one of the human obesity - related genes. Methods The cDNA of NPY2R gene was amplified from RNA of the human fat by reverse transcription polymerase chain reaction （ RT - PCR） . The PCR products were recombined with pET28a ＋ vector, positive clones were selected and DNA sequence analysis was conducted for the identification, followed by sequence analysis and homology comparison with that reported in Genebank after sequencing. Results A DNA fragment of about 1 100 bp was obtained by RT - PCR, the cDNA sequence of the recombinant plasmid of pET28a ＋ with the fragment was analyzed, and the result showed that the cloned DNA fragment was just human NPY2R eDNA. The cloned gene was encoded 381 amino acids with a molecular weight of 42 kD. The nueleotide sequence homology of the cloned NPY2R was 100% in comparison with that reported in Genebank. Conclusion The sequence of NPY2R eDNA that we have successfully cloned is the same as that of NPY2R in Genebank, which lays the foundation for further studies of gene expression of NPY2R and the interrelationship of NPY2R and the genesis, development and transformation of obesity by means of molecular biology.